KR102211477B1 - Bioink for 3d bioprinting and process for preparing the same - Google Patents

Abstract

The present invention relates to a manufacture of a new bioink for 3D bioprinting using methylcellulose, which is a temperature-sensitive polymer, and an object of the present invention is to manufacture a new bioink for 3D bioprinting that is easy to control physical properties and has excellent printability and structural stability by introducing a tyrosine group to methyl cellulose to additionally provide light-sensitive properties while maintaining temperature-sensitive properties. The physical properties of the bioink can be controlled before printing by forming a physical hydrogel based on the temperature, so that it is possible to improve printability and to manufacture the bioink with excellent structural stability by forming chemical hydrogel through photocrosslinking after printing. Vitamin derivatives serving as photoinitiators are added to natural polymers and visible light crosslinking is used, so that it is possible to manufacture a 3D structure that does not show cytotoxicity.

Description

Bioink for 3D bioprinting and its manufacturing method {BIOINK FOR 3D BIOPRINTING AND PROCESS FOR PREPARING THE SAME}

The present invention relates to a bioink for 3D bioprinting and a method of manufacturing the same.

3D printing technology can be defined as "a technology that rapidly produces 3D modeling data worked in a computer into a physical shape that can be directly touched". In the early 1980s, a 3D printer that made objects by solidifying plastic liquid Since its development, it is an industry that is constantly developing not only in automobile, aviation/space, architecture, but also fashion and medical fields.

The 3D printer has the advantage of being able to produce a small amount of various types because it is inexpensive due to a simple process and there is no limitation in the form of manufacture, but there is a limitation in that the material of bioink is large in its application to 3D bioprinting, which is a medical field.

Bio-ink is a material for 3D bioprinting in which cells, polymers, and crosslinking agents are mixed, and a variety of excellent physical properties such as biocompatibility, appropriate viscosity and viscoelastic properties are required. Synthetic polymers have an advantage in that their physical properties are easy to control and thus printability is good, but there is a problem of side effects due to low biocompatibility. In addition, since conventional natural polymers have very low physical properties, there is an inconvenience of using an additional support or mixing a thickener and another polymer to improve physical properties.

Therefore, there is currently an effort to supplement the physical properties by modifying the natural polymer and then forming a chemical hydration gel through light or enzyme crosslinking, but due to the toxicity of radiation, ultraviolet rays and crosslinking agents, the high cost of enzymes, and substrate specificity, additional 3D biotechnology There is a need to develop bioinks suitable for printing.

In order to solve the problems of the prior art as described above, the present invention introduces a tyrosine group through a two-step reaction of methylcellulose, a temperature-sensitive natural polymer, and a method for producing a natural polymer capable of double crosslinking by using a vitamin derivative as a photoinitiator. It aims to provide.

It is also an object of the present invention to provide a bioink for 3D bioprinting that is manufactured by the above manufacturing method and is easy to control physical properties, has excellent structural stability and biocompatibility.

The manufacturing method of the bio-ink for 3D bioprinting according to the present invention comprises the steps of: (a) introducing a tyrosine group into a temperature-sensitive polymer to prepare an aqueous solution of a temperature-sensitive polymer into which a tyrosine group is introduced; (b) adding a photoinitiator to the aqueous solution; Includes.

At this time, the temperature-sensitive polymer may be one selected from the group including methylcellulose (MC) and its analogs, poly (N-isopropylacrylamide), PNIPAM), and gelatin.

In addition, the step (a), the step of introducing a carboxylic acid group (-COOH) to the temperature-sensitive polymer; Substituting a tyrosine group on a carboxylic acid group; And preparing an aqueous solution by adding a temperature-sensitive polymer into which a tyrosine group is introduced into distilled water. It may include.

In addition, the photoinitiator is Riboflavin (RF), Riboflavin 5'-monophosphate sodium salt (RFp), Eosin Y (EY), Irgacure 2959 (I2959), 2,2'-Azobis[2-methyl-N-(2-hydroxyethyl) propionamide] (VA-086), Ruthenium complex (Ru), Lithium phenyl-2,4,6-trimethylbensoylphosphinate (LAP), it may be one or more selected from the group.

In addition, the present invention provides a bioink for 3D bioprinting prepared by the manufacturing method according to the present invention, wherein the bioink for 3D bioprinting may include a temperature-sensitive polymer into which a tyrosine group is introduced and a vitamin derivative as a photoinitiator. .

In addition, the present invention provides a method of manufacturing a 3D structure using the bio-ink, the method of manufacturing a 3D structure comprising the steps of: (a) introducing a tyrosine group into a temperature-sensitive polymer to prepare a temperature-sensitive polymer aqueous solution into which a tyrosine group is introduced; (b) adding a photoinitiator to the aqueous solution; (c) heating the aqueous solution to which the photoinitiator is added to a lower limit critical solution temperature; (d) printing the heated aqueous solution to prepare a structure; And (e) irradiating light to the structure. It may include.

In the present invention, by adding a vitamin derivative corresponding to the Hofmeister series to methylcellulose that has been given a photocrosslinking property through a two-step reaction, it is easy to control physical properties, improve physical properties, and improve structural stability by simultaneously expressing gelation promotion and photoinitiation properties. Bio-ink for 3D bioprinting with excellent biocompatibility can be prepared.

1 is a photograph showing a sol-gel transition phenomenon according to the temperature of an aqueous methylcellulose solution.
2 is a conceptual diagram schematically showing a mechanism for introducing a tyrosine group into methylcellulose.
3 is a conceptual diagram schematically showing a gelation mechanism of an aqueous methylcellulose solution into which a tyrosine group is introduced.
4 is a graph showing changes in gelation behavior according to the type of terminal group of methylcellulose.
5 is a conceptual diagram showing a change in gelation behavior of methylcellulose into which a tyrosine group is introduced according to the type of photoinitiator.
6 is a diagram showing the chemical structure of a photoinitiator according to the present invention.
7 is a graph showing changes in gelation behavior according to each variable of methylcellulose into which a tyrosine group is introduced.
8 is a graph showing changes in mechanical strength according to a crosslinking method of methylcellulose into which a tyrosine group is introduced.
9 is a graph showing the crosslinking density of methylcellulose into which a tyrosine group is introduced according to a crosslinking method.
10 is a photograph showing the structure of a 3D structure made with bio-ink according to the present invention.
11 is a photograph and graph showing the biocompatibility of the 3D structure made with the bio-ink according to the present invention.

In the present invention, various modifications may be made and various forms may be applied, and specific embodiments will be illustrated in the drawings and described in detail in the text. However, this is not intended to limit the present invention to a specific form disclosed, it should be understood to include all changes, equivalents, and substitutes included in the spirit and scope of the present invention.

In the present application, terms such as "comprise" or "have" are intended to designate the presence of features, numbers, steps, actions, components, parts, or combinations thereof described in the specification, but one or more other features. It is to be understood that the presence or addition of elements or numbers, steps, actions, components, parts, or combinations thereof, does not preclude in advance the possibility of being added. Further, when a part such as a layer, film, region, plate, etc. is said to be "on" another part, this includes not only the case where the other part is "directly above", but also the case where there is another part in the middle. Conversely, when a part such as a layer, film, region, plate, etc. is said to be "under" another part, this includes not only the case where the other part is "directly below", but also the case where there is another part in the middle. In addition, in the present application, the term "above" may include a case where it is disposed not only above but also below.

Hereinafter, the present invention will be described in detail.

The manufacturing method of the bio-ink for 3D bioprinting according to the present invention comprises the steps of: (a) introducing a tyrosine group into a temperature-sensitive polymer to prepare an aqueous solution of a temperature-sensitive polymer into which a tyrosine group is introduced; (b) adding a photoinitiator to the aqueous solution; Includes.

That is, it is possible to provide a bio-ink for 3D bioprinting that is easy to control physical properties including a natural polymer and a photoinitiator that can be double-crosslinked and has excellent biocompatibility, prepared through a two-step reaction.

The temperature-sensitive polymer may be one selected from the group including methylcellulose (MC) and its analogs, poly (N-isopropylacrylamide), PNIPAM), and gelatin.

The temperature-sensitive polymer is a polymer having a lower critical solution temperature (LCST) rather than an upper critical solution temperature (UCST), and methylcellulose is most preferred in terms of double gelation behavior.

로 증가함에 따라 분자 내/간 소수성 상호작용의 증가로 젤화가 일어나는 것을 확인할 수 있다.Methylcellulose is a representative temperature-sensitive cellulose derivative. As the temperature increases, the intramolecular/intermolecular hydrophobic interaction increases, and a sol-gel transition occurs above a certain temperature. These temperature-sensitive properties can be controlled by the concentration, molecular weight, and additives of the polymer, and in the case of additives, the gelation is accelerated by promoting the salting out effect in the solution. 1 shows the gelation behavior according to the temperature of a low molecular weight methylcellulose aqueous solution, and maintains the solution state at room temperature, but the temperature is 37

It can be seen that gelation occurs due to an increase in intramolecular/intermolecular hydrophobic interactions as it increases.

When the temperature-sensitive polymer is methylcellulose, the degree of substitution (DS) of the methylcellulose is preferably 1.5 to 1.8, and more preferably 1.6 to 1.7. When the degree of substitution of methylcellulose is within the above range, it is well dissolved in cold water. In addition, the viscosity of the low molecular weight methylcellulose is preferably 10 to 50 cP, more preferably 15 to 20 cP. In addition, the molecular weight of the low molecular weight methylcellulose is preferably 50,000 to 200,000 Dalton, more preferably 100,000 to 150,000 Dalton. If the molecular weight of low molecular weight methylcellulose exceeds 200,000 Daltons, the solubility of high molecular weight methylcellulose is less than 3% by weight, which is not preferable.

2 is a conceptual diagram schematically showing a mechanism for introducing a tyrosine group into methylcellulose.

Referring to Figure 2, the step (a), the step of introducing a carboxylic acid group (-COOH) to the temperature-sensitive polymer (eg, methylcellulose); Substituting a tyrosine group on a carboxylic acid group; And preparing an aqueous solution by adding a temperature-sensitive polymer into which a tyrosine group is introduced into distilled water. It may include. That is, in the present invention, the step of introducing a tyrosine group to a temperature-sensitive polymer, preferably methylcellulose, may consist of a one-step reaction of introducing a carboxylic acid group (-COOH) and a two-step reaction of substituting a tyrosine group to a carboxylic acid group.

In the first step reaction, it is preferable to add succinic acid (SA) and triethylamine (TEA) to the temperature-sensitive polymer in a molar ratio of 1:0.01 to 100, and to add a molar ratio of 1:0.4 to 4. More preferable. The reaction time is preferably 1 to 120 hours, more preferably 12 to 24 hours.

In the two-step reaction, carbodiimide (1-Ethyl-3-(3-dimethylaminopropyl)-carbodiimide, EDC), hydroxysuccinimide (NHS), and tyrosine-introduced materials were added to a temperature-sensitive polymer into which a carboxylic acid group was introduced. As an addition reaction, the carbodiimide and hydroxysuccinimide are preferably added in a molar ratio of 1:0.01 to 100 per number of repeating units of the polymer, and more preferably in a molar ratio of 1:1 to 10. The reaction time is preferably 5 minutes to 120 hours, more preferably 1 to 12 hours. Thereafter, the tyrosine-introducing material is preferably added in a molar ratio of 1:0.01 to 100 per number of repeating units of the polymer, and more preferably in a molar ratio of 1:1 to 10. The reaction time is preferably 1 to 120 hours, more preferably 12 to 48 hours. The tyrosine introduction material may be one selected from the group containing tyramine, dopamine, hydroxyphenylacetic acid, hydroxypropionic acid, epinephrine, and hydroxyethylaniline, and tyramine is preferred.

When a tyrosine group is introduced into the temperature-sensitive polymer, a powdery temperature-sensitive polymer is added to distilled water and stirred at room temperature to prepare an aqueous solution. The content of the temperature-sensitive polymer in the aqueous solution is preferably 1 to 15% by weight, more preferably 8 to 10% by weight.

When a temperature-sensitive polymer aqueous solution into which a tyrosine group is introduced is prepared, a photoinitiator may be added to the aqueous solution.

In this case, as photoinitiators, Riboflavin (RF), Riboflavin 5'-monophosphate sodium salt (RFp), Eosin Y (EY), Irgacure 2959 (I2959), 2,2'-Azobis[2-methyl-N-(2-hydroxyethyl) )propionamide] (VA-086), Ruthenium complex (Ru), Lithium phenyl-2,4,6-trimethylbensoylphosphinate (LAP), and at least one selected from the group containing Riboflavin (RF), riboflavin 5' -monophosphate sodium salt (RFp) is preferred. That is, by using a vitamin derivative as a photoinitiator, bio-inks with excellent biocompatibility can be prepared, and by promoting the salting out effect and hydrophobic interactions between temperature-sensitive polymers, the gelation rate and gel strength are improved, and photocrosslinking is possible in the visible light region. Let's do it.

In addition, in addition to the photoinitiator, a co-initiator may be further added to the temperature-sensitive polymer aqueous solution. At this time, the open initiator is selected from the group including arginine (2-Amino-5-guanidinopentanoic acid, L-arginine), Dimethylaminoethyl acrylate (DMAEMA), Neopenthyl Glycol (NPG), and sodium persulfate (SPS). More than one type may be used, and sodium persulfate (SPS) is preferred.

Sodium persulfate (SPS), which is a co-agent, is preferably added to 1 to 500 mM, more preferably 10 to 200 mM. The photoinitiators riboflavin (RF) and riboflavin 5'-monophosphate sodium salt (RFp) are preferably 0.01 to 110 mM, and more preferably 0.05 to 10 mM.

In addition, the present invention provides a method of manufacturing a 3D structure using the bio-ink, the method of manufacturing a 3D structure comprising the steps of: (a) introducing a tyrosine group into a temperature-sensitive polymer to prepare a temperature-sensitive polymer aqueous solution into which a tyrosine group is introduced; (b) adding a photoinitiator to the aqueous solution; (c) heating the aqueous solution to which the photoinitiator is added to a lower limit critical solution temperature; (d) printing the heated aqueous solution to prepare a structure; And (e) irradiating light to the structure. It may include.

That is, the physical hydration gel for the bioink for 3D bioprinting according to the present invention is formed by temperature to increase printability by controlling the physical properties of the bioink before printing, and structural stability by forming a chemical hydrogel through photocrosslinking after printing. Can manufacture excellent 3D structures.

3 is a conceptual diagram schematically showing a gelation mechanism of an aqueous methylcellulose solution into which a tyrosine group is introduced.

Referring to FIG. 3, when a temperature higher than the lower critical solution temperature (LCST) is applied to an aqueous methylcellulose derivative having both temperature-sensitive and light-sensitive properties, a physical hydrogel due to physical crosslinking of hydrophobic interactions is formed. And additionally irradiated with light to form a chemical hydrogel due to chemical crosslinking of tyrosine groups. At this time, photo-crosslinking can be formed by adding a co-initiator and a photo-initiator to the methylcellulose aqueous solution into which the tyrosine group is introduced.

The optical crosslinking time is preferably 10 to 600 seconds, more preferably 30 to 360 seconds. The wavelength band is preferably 200 to 800 nm, more preferably 380 to 780 nm, and even more preferably 400 to 500 nm (visible light).

In addition, the present invention provides a bioink for 3D bioprinting prepared by the manufacturing method according to the present invention, wherein the bioink for 3D bioprinting may include a temperature-sensitive polymer into which a tyrosine group is introduced and a vitamin derivative as a photoinitiator. have. That is, in the bioink, a temperature-sensitive polymer into which a tyrosine group is introduced is double-crosslinked by heat and light, and a vitamin derivative is used as a photoinitiator, so that a 3D structure with excellent structural stability, non-toxicity and excellent biocompatibility can be produced. I can.

Hereinafter, the present invention will be described in detail through examples. However, the following examples are only illustrative of the present invention, and the present invention is not limited by the following examples.

[Example 1]

Preparation of temperature-sensitive polymer with tyrosine group introduced

As a temperature-sensitive polymer, succinic acid (SA) and triethylamine (TEA) were added to methylcellulose (substitution degree 1.6-1.7, viscosity 15 cP, molecular weight 110,000 Dalton) in a molar ratio of 1:0.4, and 12 hours During the reaction, a carboxylic acid group was introduced into methylcellulose.

Carbodiimide (1-Ethyl-3-(3-dimethylaminopropyl)-carbodiimide, EDC) and hydroxysuccinimide (NHS) were added to the methylcellulose into which the carboxylic acid group was introduced in a molar ratio of 1:1.8. After reacting for 1 hour, tyramine (Tyr) was added to the reactant at a molar ratio of 1:0.9, and the reaction was carried out for 24 hours to prepare a temperature-sensitive polymer into which a tyrosine group was introduced.

Preparation of a temperature-sensitive polymer aqueous solution into which tyrosine groups are introduced

A methylcellulose derivative in powder form was added to distilled water and stirred at room temperature for 24 hours to prepare an 8% by weight aqueous methylcellulose solution.

Gelation of temperature-sensitive polymer aqueous solution with tyrosine group introduced

After adding 100 mM sodium persulfate (SPS) as a co-initiator and 0.25 mM Riboflavin (RF) and Riboflavin 5'-monophosphate sodium salt (RFp) as a photoinitiator to the methylcellulose aqueous solution, the aqueous solution was added to the lower critical solution temperature. solution temperature, LCST) or higher. Thereafter, photocrosslinking was performed by irradiating visible light having a wavelength of 400-500nm for 120 seconds to obtain methylcellulose into which a double-crosslinked tyrosine group was introduced.

[Experimental Example 1]

의 가열속도로 측정되었고 젤화시간은 37

조건 하에서 측정되었다. 그 결과는 도 4에 나타내었다.The gelation behavior according to the type of terminal group of the methylcellulose derivative was measured using a rheometer. At this time, the strain and frequency were 0.3% and 1 rad/s, and the gelation temperature was 1

It was measured at the heating rate of and the gelation time was 37

Measured under conditions. The results are shown in FIG. 4.

Referring to FIG. 4, the gelation temperature and time of the methylcellulose derivative confirmed that gelation was suppressed by the terminal group, and the suppression of gelation by the tyrosine group was greater than that of the carboxylic acid group (-COOH), which resulted in a larger terminal group. I could see that it was because it became hard. However, even though the gelling behavior was slowed according to the type of terminal group, it was confirmed that gelation still occurred near body temperature, and thus it was confirmed that the double-crosslinked methylcellulose prepared in the present invention still has a temperature-sensitive characteristic.

[Experimental Example 2]

의 가열속도로 측정되었고 젤화시간은 37

조건 하에서 측정되었다. G'과 G"의 교차점은 젤화지점을 나타내며 광개시제인 비타민유도체의 화학구조식은 도 6에 제시하였다.The gelation behavior of the double-crosslinked methylcellulose according to the photoinitiator type was measured using a rheometer. At this time, the strain and frequency were 0.3% and 1 rad/s, and the gelation temperature was 1

It was measured at the heating rate of and the gelation time was 37

Measured under conditions. The intersection of G'and G" represents the gelation point, and the chemical structural formula of the vitamin derivative as a photoinitiator is shown in FIG.

Referring to FIG. 5, it was confirmed that gelation temperature and time were accelerated when a phosphate group was included in the vitamin derivative as shown in FIG. 6(B), and this was found to be due to the salt effect by the Hofmeister series. In this way, if the vitamin derivative contains a phosphate group, gelation is promoted because it attracts water from the aqueous solution and promotes the hydrophobic interaction of the hydrophobic group of methylcellulose. Using this phenomenon, the formation behavior of the physical hydrogel can be easily controlled. It was confirmed that the resulting gelation delay can be compensated.

[Experimental Example 3]

조건 하에서 젤화시간이 측정되었다. The gelation behavior of methylcellulose into which the tyrosine group was introduced was measured using a rheometer. At this time, the strain and frequency were 0.3% and 1 rad/s, and 37

The gelation time was measured under conditions.

According to FIG. 7, it was confirmed that gelation was promoted as the concentration of the open initiator, the concentration of the photoinitiator, and the light irradiation time increased, and the gel strength also increased. It was confirmed that the physical properties of the chemical hydrogel can be easily controlled by adjusting these various variables.

[Experimental Example 4]

8 is a graph showing changes in mechanical strength according to a crosslinking method of methylcellulose into which a tyrosine group is introduced.

Referring to FIG. 8, compared to the case of forming the hydration gel by physical and chemical single crosslinking, the hydration gel subjected to double crosslinking was significantly improved in physical properties, and vitamin derivatives were added as photoinitiators according to the type of photoinitiator. It was confirmed that the mechanical strength was higher when it was done.

[Experimental Example 5]

9 is a graph showing the crosslinking density of methylcellulose into which a tyrosine group is introduced according to a crosslinking method.

Referring to FIG. 9, it can be seen that the light efficiency is very good as it has a crosslinking density of about 80% in the case of double crosslinking. Since the introduced tyramine is only 2.6% of the total repeating unit, there is no significant effect, but the light efficiency is also very good, and since there are not many unphotocrosslinked tyramine groups remaining after crosslinking and printing, it is excellent in biocompatibility and there are no side reactions.

[Experimental Example 6]

Various shapes were prepared using the previously prepared double-crosslinked methylcellulose-based ink. Specifically, two types of 3D structures (Preparation Example 1 and Preparation Example 2) were produced, and FIG. 10 is a photograph showing the structure of a 3D structure made with bio-ink according to the present invention. Referring to FIG. 10, (a) of FIG. 10 is a photograph showing the structure of Preparation Example 1, and (b) is a photograph showing the structure of Preparation Example 2.

Referring to FIG. 10, it was confirmed that ink having good printability was prepared regardless of the shape and size. At this time, the optimum conditions were established through analysis of various variables (temperature, needle diameter, speed, extrusion volume, etc.), and a double-crosslinked methylcellulose aqueous solution was added to the syringe, and the viscosity and viscosity suitable for extrusion-based printing before printing. By applying temperature so as to have surface tension, a physical hydrogel was formed and then printed to produce a structure with excellent surface characteristics of the printed filaments or interactions between filaments, and by irradiating visible light while printing the filaments. A 3D structure with excellent structural stability could be obtained.

[Experimental Example 7]

The biocompatibility of the 3D structure prepared in Experimental Example 6 was confirmed. The results are shown in FIG. 11.

Referring to FIG. 11, (a) is a result of measuring the biocompatibility of Preparation Example 1, and (b) is a result of measuring the biocompatibility of Preparation Example 2, and both Preparation Example 1 and Preparation Example 2 have high cell viability. As the time passed from 4 hours to 72 hours, the number of cells increased, confirming that proliferation occurred.

Although the above has been described with reference to a preferred embodiment of the present invention, those skilled in the art or those of ordinary skill in the art will not depart from the spirit and scope of the invention described in the claims to be described later. It will be understood that various modifications and changes can be made to the present invention within the scope of the invention.

Therefore, the technical scope of the present invention should not be limited to the content described in the detailed description of the specification, but should be determined by the claims.

Claims (6)

(a) preparing an aqueous solution of a temperature-sensitive polymer into which a tyrosine group is introduced by introducing a tyrosine group into the temperature-sensitive polymer;
(b) adding a photoinitiator to the aqueous solution; Including,
The temperature-sensitive polymer is one selected from the group including methylcellulose (MC) and its analogues, poly (N-isopropylacrylamide), PNIPAM), and gelatin for 3D bioprinting. Method for producing bio-ink.
delete The method of claim 1,
The step (a),
Introducing a carboxylic acid group (-COOH) to the temperature-sensitive polymer;
Substituting a tyrosine group on a carboxylic acid group; And
Preparing an aqueous solution by introducing a temperature-sensitive polymer into which a tyrosine group is introduced into distilled water; Method for producing a bio-ink for 3D bioprinting, characterized in that it comprises a.
The method of claim 1,
The photoinitiator is Riboflavin (RF), Riboflavin 5'-monophosphate sodium salt (RFp), Eosin Y (EY), Irgacure 2959 (I2959), 2,2'-Azobis[2-methyl-N-(2-hydroxyethyl)propionamide ](VA-086), Ruthenium complex (Ru), Lithium phenyl-2,4,6-trimethylbensoylphosphinate (LAP) A method for producing a bio-ink for 3D bioprinting, characterized in that at least one selected from the group containing.
(a) preparing an aqueous solution of a temperature-sensitive polymer into which a tyrosine group is introduced by introducing a tyrosine group into the temperature-sensitive polymer;
(b) adding a photoinitiator to the aqueous solution;
(c) heating the aqueous solution to which the photoinitiator is added to a lower limit critical solution temperature;
(d) printing the heated aqueous solution to prepare a structure; And
(e) irradiating light to the structure; Including,
The temperature-sensitive polymer is methylcellulose (MC) and its analogues, poly(N-isopropylacrylamide), PNIPAM), and gelatin (gelatin). Way.
A bioink for 3D bioprinting prepared by the manufacturing method according to any one of claims 1, 3 and 4, comprising a temperature-sensitive polymer into which a tyrosine group is introduced and a vitamin derivative as a photoinitiator,
The temperature-sensitive polymer is one selected from the group including methylcellulose (MC) and its analogues, poly (N-isopropylacrylamide), PNIPAM), and gelatin for 3D bioprinting. Bioink.

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